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Hypocalcemia, defined as total blood calcium concentrations below 2.1 mM, has detrimental impacts on welfare, production and reproduction in dairy cattle. Yet, no cow-side test exists for testing total bovine blood calcium. Here, we modified the split trehalase complementation assay to detect total calcium in serum by incorporating calmodulin and the M13 peptide as fusion partners to the trehalase fragments. In the presence of calcium, calmodulin undergoes a conformation change and gains strong affinity for M13 peptide. A high reactive assay for calcium was developed with detection threshold of 10 uM and dynamic range between 1 uM and 1 mM. The addition of a specific concentration of calcium chelator, EDTA, in mild acidic conditions, shifted the dynamic range to physiological calcium concentrations and transformed the sensor from ionized calcium sensor to total calcium sensor. The sensor was validated on a collection of 213 bovine serum samples by comparison with quantitative colorimetric calcium test. A correlation coefficient of 0.81 was achieved and the accuracy of detecting subclinical hypocalcemia was 0.86 and specificity of 100%. The area under the Receiver Operating Characteristic curve was 0.93. The concordance correlation coefficient (0.80), Bland-Altman plot and weighted Kappa coefficient (0.71) demonstrated a substantial agreement between both methods. In conclusion, a novel total calcium test was developed that can be used as a convenient high throughput laboratory test and with potential to be incorporated into a version compatible with on-farm testing., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Abstract: Milk fever is an important metabolic disorder of dairy cows after calving, and is characterized by hypocalcemia, tetany, lateral recumbency, and eventual coma. To date, there have been many reports about the pathogenesis and pathophysiology of milk fever, but the plasma protein profile in milk fever has not been reported. The aim of our study was to investigate novel pathophysiological changes in the plasma proteome of cows affected with milk fever. Plasma samples were collected from eight Holstein cows with milk fever (T), and eight control Holstein cows without milk fever (C), at an intensive Holstein dairy farm in Heilongjiang province, China. Samples were analyzed by fluorescence two-dimensional (2D) differential in-gel electrophoresis (DIGE), followed by in-gel digestion, and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF-MS) for peptide mass fingerprinting of selected protein spots. Eight of the 23 differential protein spots in the plasma of T and C cows were isolated and identified by 2D-DIGE and MALDI-TOF-MS. The protein spots represented five unique proteins, and had significant alterations in spot volume as determined by DeCyder differential in-gel analysis (DIA) software. The upregulated proteins were identified as serpin peptidase inhibitor (angiotensin), which regulates blood pressure and maintains fluid and electrolyte homeostasis, and endopin 2B which is involved in neural regulation. The downregulated proteins were serum albumin, which acts as a transport protein, fibrinogen beta chain which is involved in blood coagulation, and IgG heavy-chain C-region (IgG-CH) which participates in the immune response. In conclusion, we were able to use proteomic technologies to identify several novel plasma proteins in cows affected with milk fever. These findings may reveal new pathophysiological changes that occur in cows with milk fever. [Copyright &y& Elsevier]

Abstract: The objective of this study was to estimate heritability and genetic correlations between the liabilities of clinical mastitis (CM), milk fever (MF), metritis (MET), and retained placenta (RP) within the first three lactations of Holstein dairy cows. The records of 57,301 dairy cows from 20 large dairy herds in Iran between January 2005 and June 2009 were analysed with univariate and bivariate threshold animal models, using Gibbs sampling methodology. The final model included the fixed class effects of herd-year, season of calving, parity of dam, the linear covariate effect of age at calving, and the random direct genetic effect of animal. Posterior means of heritability for liabilities in first, second, and third lactations were 0.06, 0.08, and 0.09, respectively, for CM; 0.10, 0.12, and 0.11, respectively, for MF; 0.09, 0.07, and 0.10, respectively, for MET, and 0.07, 0.08, and 0.08, respectively, for RP. Posterior means of genetic correlations between disease liabilities were low or moderate (from −0.01 to 0.26). The results of this study indicated the importance of health traits for considering in the selection index of Iranian Holstein dairy cows. [ABSTRACT FROM AUTHOR]

Abstract: The objectives of this study were to determine the effects of a pre-partum diet with lower than recommended (DCAD=−82mEq/kg of dietary DM) amounts of anionic salts on metabolism, health, reproductive performance and milk production in dairy cows. Sixty Holstein multiparous cows were enrolled 21days prior to expected calving date. The animals were randomly assigned to receive one of two rations: 30 cows received anionic ration [−82mEq (NA+K−Cl−S)/kg of DM] for 21d to parturition and the other group (n =30) were fed a usual dry period ration (+192mEq/kg of DM). Serum samples obtained at days −21, +3 and +21 relative to calving were analyzed for β-hydroxybutyrate (BHBA), non-esterified fatty acids (NEFA), glucose, calcium (Ca), inorganic phosphorus, magnesium, chloride, sodium, potassium, cholesterol, urea, creatinine, total protein, albumin, and aspartate aminotransferase (AST). Urine pH declined from 8.4 at 21d before calving (pre-treatment) to 6.2 at day 7 pre-partum in the treatment group. Repeated-measure mixed model analysis indicated that the concentrations of Ca were significantly increased and creatinine, and AST were significantly decreased by lowering DCAD. The concentrations of BHBA, NEFA and glucose were not affected by treatment. The incidence of milk fever and culling were 5 and 11 times higher in the control group in comparison with the treatment group, respectively. The intervals from calving to first breeding and to pregnancy were not influenced by treatment. There was no group effect on average daily milk yield or fat percentage. [Copyright &y& Elsevier]

Providing tablets of 1,25-dihydroxyvitamin D 3 (1,25(OH) 2 D 3 ), the biologically active metabolite of vitamin D 3 , in a rumen bolus may be used as prevention for periparturient hypocalcemia in dairy cows. This study investigated the pharmacokinetics of 1,25(OH) 2 D 3 glycosides extracted from Solanum glaucophyllum (SGE) on blood serum 1,25(OH) 2 D 3 , Ca, P and Mg response in dry pregnant dairy cows. Boluses contained tablets of SGE which differed in their release properties (rapid release, slow release and combination) and galenics (200 μg uncoated, 300 μg and 500 μg uncoated or coated, 2 × 500 μg uncoated). Nineteen blood samples were collected from 29 cows between 96 h before and 336 h after bolus administration. Blood serum 1,25(OH) 2 D 3 , Ca and P increased between 12 h and 120 h, 12 h and 264 h and 24 h and 264 h, respectively. Highest values were reached at 30 h, 72 h and 120 h for 1,25(OH) 2 D 3 , Ca and P, respectively. Baseline values were then reached at 216 h for 1,25(OH) 2 D 3 and 336 h for Ca and P. Concentration of Mg decreased between 24 h and 216 h, before reaching values comparable to baseline at 264 h. Highest Ca values were obtained with the combined rapid and slow release properties (500 μg) and there was no effect from coating on pharmacokinetics. In conclusion, the antepartum oral SGE bolus administration may be suitable for the prevention of periparturient hypocalcemia., (Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Osteoprotegerin (OPG) inhibits osteoclast (OC) differentiation. TRAP5b (tartrate-resistant acid phosphatase 5b) secreted by OCs reflects the numbers of mature OCs. This study assessed these OC-related markers around parturition in cows of different parities and in cows with milk fever (MF). The blood OPG and TRAP5b concentrations, as well as the ratio of OPG to TRAP5b (O/T), were measured beginning 3 weeks before (-21 d) and over a few days after calving in 49 Holstein Friesian cows at first (n = 8), second (n = 17), third (n = 12), and fourth or greater (n = 12) parities. The ratio of O/T at -21 d to O/T at calving (PreCOT) was also calculated. In the third and greater parities, seven cows developed MF (non-MF, n = 17). Regardless of the development of MF, the serum OPG started to decline in the last week of gestation only in the cows entering the second lactation, while the blood TRAP5b increased at calving in all cows. O/T decreased toward parturition only in multiparous cows. The decrease in O/T at caving was less pronounced in MF cows. PreCOT was negatively correlated with lactation number only in multiparous cows (n = 41, ρ = -0.50, P < .01). This study implied that OC differentiation toward calving was tapered in cows with advanced parities, and these indexes predict the risk of MF., Competing Interests: Declaration of Competing Interest The authors declare no conflicts on interests., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

The objective of this study was to search for potential alterations in innate immunity reactants and carbohydrate and lipid metabolism in the blood of transition dairy cows before, during, and after clinical occurrence of milk fever (MF) and identify potential predictive biomarkers of disease. One hundred pregnant multiparous Holstein dairy cows were involved in the study starting from -8wks before the expected day of parturition until +8wks postpartum as part of a large retrospective longitudinal study. Health status, DMI, milk yield, and milk composition were monitored during the whole experimental period. Six healthy cows (CON) and 6 cows that showed clinical signs of MF were selected for blood analyses. Serum concentrations of lactate, non-esterified fatty acids (NEFA), β-hydroxybutyric acid (BHBA), interleukin-1 (IL-1), interleukin-6 (IL-6), tumor necrosis factor (TNF), haptoglobin (Hp), and serum amyloid A (SAA) were determined. Results indicated that concentrations of serum lactate, IL-6, TNF, SAA, and Hp were greater in cows with MF than those in the CON group at different time points. Moreover, serum lactate, TNF, SAA, and Hp were greater in cows with MF starting at -8 and -4wks prior to parturition. Both principal component analysis (PCA) and partial least squares - discriminant analysis (PLS-DA) showed separated clusters between MF and CON cows at -8, -4, and disease diagnosis weeks. Overall DMI and milk production were lower in MF-affected cows. Additionally milk fat and fat:protein ratio were greater in MF. In conclusion, cows affected by MF showed alterations in some of the innate immunity reactants and metabolites related to carbohydrate metabolism several weeks prior to appearance of clinical signs of MF. Variable importance in projection plots demonstrated that TNF and SAA in the serum were the strongest discriminators between MF cows and CON ones, which might be useful as predictive biomarkers of the disease., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

The objective of this study was to search for potential alterations in innate immunity reactants and carbohydrate and lipid metabolism in the blood of transition dairy cows before, during, and after clinical occurrence of milk fever (MF) and identify potential predictive biomarkers of disease. One hundred pregnant multiparous Holstein dairy cows were involved in the study starting from -8wks before the expected day of parturition until +8wks postpartum as part of a large retrospective longitudinal study. Health status, DMI, milk yield, and milk composition were monitored during the whole experimental period. Six healthy cows (CON) and 6 cows that showed clinical signs of MF were selected for blood analyses. Serum concentrations of lactate, non-esterified fatty acids (NEFA), β-hydroxybutyric acid (BHBA), interleukin-1 (IL-1), interleukin-6 (IL-6), tumor necrosis factor (TNF), haptoglobin (Hp), and serum amyloid A (SAA) were determined. Results indicated that concentrations of serum lactate, IL-6, TNF, SAA, and Hp were greater in cows with MF than those in the CON group at different time points. Moreover, serum lactate, TNF, SAA, and Hp were greater in cows with MF starting at -8 and -4wks prior to parturition. Both principal component analysis (PCA) and partial least squares - discriminant analysis (PLS-DA) showed separated clusters between MF and CON cows at -8, -4, and disease diagnosis weeks. Overall DMI and milk production were lower in MF-affected cows. Additionally milk fat and fat:protein ratio were greater in MF. In conclusion, cows affected by MF showed alterations in some of the innate immunity reactants and metabolites related to carbohydrate metabolism several weeks prior to appearance of clinical signs of MF. Variable importance in projection plots demonstrated that TNF and SAA in the serum were the strongest discriminators between MF cows and CON ones, which might be useful as predictive biomarkers of the disease., (Copyright © 2017 Elsevier Ltd. All rights reserved.)